Vibrio vulnificus (Vv), an emerging Gram-negative bacterial pathogen, is found in warm marine environments worldwide. Chronic liver disease, particularly alcohol-induced liver disease, is a major risk factor for severe Vv infection. Even with antibiotic treatment, the mortality rate for severe Vv infection can exceed 50%. Most fatalities are caused by septic shock that results from dysregulation of proinflammatory cytokines such as TNF1, presumably due to interaction of Vv agonists with Toll-like receptors (TLRs). TLRs are a family of pattern recognition receptors that play a major role in the harmful inflammatory response. A recent study showed that Vv lipoprotein elicits TNF1 production by human monocytes via TLR2 signaling. Our preliminary data obtained with Vv ATCC 27562, a clinical (blood) isolate, demonstrates that: (1) TLR4 signaling plays a key role in TNF1 production by mouse blood and splenocytes stimulated with Vv cells;(2) TLR4 signaling is deleterious in a mouse model of Vv infection;(3) signaling by TLR(s), exclusive of TLR4, is needed to eradicate Vv infection;and (4) the TLR-mediated TNF1 response plays a critical role in determining the outcome of Vv infection. The high morbidity and mortality associated with Vv-induced sepsis underscore the need for new therapeutic strategies. We hypothesize that selective blockade of the TLR-mediated harmful inflammatory response with TLR antagonist(s), in combination with antibiotic therapy, will improve the clinical outcome of patients with Vv-induced sepsis. The long-term goal of this study is to gain a better understanding of the TLR-mediated host response to Vv in order to develop new adjunctive therapies that block harmful TLR signaling. Specific aims are to: (1) Compare, using ELISA, the TNF1 response of wild-type (WT), TLR2 knockout (KO), TLR4 KO and TLR2/TLR4 KO mouse whole blood and resident liver macrophages stimulated ex vivo with the genotypically distinct Vv clinical strains ATCC 27562, MO6-24, YJ016, and C7184;(2) Determine, using the mouse sepsis model, the effect of TLR2 KO and TLR2/TLR4 KO on the ability of Vv-infected mice to survive and clear infection;and (3) Investigate, using the mouse sepsis model, the effect of selective blockade of TLR signaling combined with antibiotic therapy on the ability of Vv-infected WT control mice and WT mice with alcohol-induced liver disease to survive and clear infection. This study will yield new information that is essential for designing novel therapeutic strategies for serious infections caused by Vv and potentially other pathogens that damage host tissues via the TLR-mediated inflammatory response. PUBLIC HEALTH RELEVANCE: Morbidity and mortality rates are high for patients with severe V. vulnificus (Vv) infection, particularly those with alcohol-induced liver damage. This study will provide new information that will (a) significantly enhance our limited understanding of the role of TLRs in the innate response to Vv and (b) guide development of new adjunctive therapies to improve the clinical outcome of patients with Vv-induced sepsis by selectively blocking TLR signaling that mediates the harmful inflammatory response. This study will also provide basic information that is relevant to the design of new therapeutic strategies for treatment of sepsis caused by other pathogenic microbes that evoke a harmful TLR-mediated inflammatory response.